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Biomedical subjects

R F Gagel

Publications and source records attributed to R F Gagel.

At least 109 records · Page 6Linked to original sources
PubMed1988

Risk estimation and screening in families of patients with medullary thyroid carcinoma.

Many gene carriers for multiple endocrine neoplasia type 2a (MEN 2a) do not manifest the disease, even into old age. Thus, a negative family history in a patient presenting with medullary thyroid carcinoma does not reliably exclude familial disease. Data are reported for the probability that the MEN 2a gene will either have manifested clinically or be detectable by stimulated calcitonin screening by a given age. These probabilities can be used to calculate individual risks and to guide screening.

Adolescent↗
PubMed1988

Synthetic human parathyroid hormone-(1-34) for the study of pseudohypoparathyroidism.

The synthetic amino-terminal fragment of PTH, PTH-(1-34), was recently released for clinical testing of PTH responsiveness. We measured the urinary cAMP and phosphaturic responses to infusion of PTH-(1-34) [3U/kg BW (200 U maximum), iv in 10 min] in patients with pseudohypoparathyroidism and idiopathic hypoparathyroidism, as well as normal subjects. The protocol used data from 5 30-min urine collections and 4 blood samples. Based on the results in 7 patients with pseudohypoparathyroidism (hypocalcemia with increased serum immunoreactive PTH concentrations), 2 patients with suspected pseudohypoparathyroidism, 9 patients with surgical hypoparathyroidism, and 10 normal subjects, this testing protocol differentiated well among these conditions. The patients with pseudohypoparathyroidism had blunted cAMP and phosphaturic responses to PTH-(1-34) administration compared to those of either normal or hypoparathyroid subjects. Induced hypercalcemia failed to restore a normal cAMP response to PTH-(1-34) infusion in 2 patients with pseudohypoparathyroidism. Calculation of the cAMP response to PTH-(1-34) as nanomoles per dL glomerular filtrate during the first 30 min after infusion provided better differentiation among groups than other parameters of cAMP metabolism. Calculating the phosphaturic response as the percent fall in tubular maximum for phosphate reabsorption during the first hour after infusion gave the best degree of statistical separation among groups. We conclude that this new diagnostic agent is effective for the study of renal responsiveness to PTH, and that the protocol described here reliably differentiates patients with pseudohypoparathyroidism from those with hypocalcemia due to other causes.

Adult↗
PubMed1988

Treatment of Paget's disease of bone with salmon calcitonin nasal spray.

Subcutaneous daily or bidaily administration of synthetic salmon calcitonin is an effective form of therapy for Paget's disease, but the requirement for parenteral injection deters geriatric patients from using the drug. This study compares a new intranasal preparation of salmon calcitonin to subcutaneous drug in 18 patients with Paget's disease using two different protocols. In the first protocol, 15 patients not previously treated with salmon calcitonin were given the agent for 3 months by either the intranasal or subcutaneous route. Seven patients treated with intranasal calcitonin had a mean fall in the serum alkaline phosphatase of 33% over a 3-month period compared to a fall of 40% in the subcutaneously treated group; the difference between the two treatment groups was not statistically significant. In the second protocol, three patients previously stabilized on subcutaneous calcitonin were switched to the nasal spray with no subsequent change in alkaline phosphatase values during 6 months of treatment. These results demonstrate that intranasal salmon calcitonin is effective in lowering the serum alkaline phosphatase in Paget's disease. Ease of administration and patient acceptance make intranasal calcitonin a reasonable alternative for geriatric patients.

Administration, Intranasal↗
PubMed1987

The effect of 1,25-dihydroxyvitamin D3 treatment on calcitonin and calcitonin gene-related peptide mRNA levels in cultured human thyroid C-cells.

We utilized the TT cell, a human C-cell line derived from a medullary thyroid carcinoma, to study the effects of 1,25-dihydroxyvitamin D3 on cell growth and expression of the calcitonin gene. The growth rate of cells treated for 8 days with 1,25-dihydroxyvitamin D3 did not differ significantly from control or 24,25-dihydroxyvitamin D3 treated cells. Total RNA was isolated, and calcitonin and calcitonin gene-related peptide (CGRP) levels were measured by hybridization. 1,25 D3 lowered calcitonin and CGRP mRNA levels in a time- and dose-dependent fashion; 24,25 D3 had no effect. Northern blots revealed a decrease in the mature mRNA as well as the common precursor forms, indicating a transcriptional effect of 1,25 D3.

24,25-Dihydroxyvitamin D 3↗
PubMed1987

Studies of short-term secretion of peptides produced by alternative RNA processing.

Alternative RNA processing of the calcitonin gene primary transcript results in production of two peptides, calcitonin and calcitonin gene-related peptide (CGRP). We have used the TT cell line, which produces both peptides, to ascertain whether secretion of peptides produced by alternative RNA processing is under identical regulatory control. Short-term treatment of TT cells with phorbol esters and cAMP analogs caused a rapid and parallel release of both calcitonin and CGRP. The measurement of calcitonin and CGRP mRNA levels during treatment revealed that new RNA synthesis was not required for secretion. Four potential regulators of phorbol ester-mediated and five of cAMP-mediated secretion were identified by incorporation of radioactive phosphate into protein as analysed by two-dimensional polyacrylamide gel electrophoresis and autoradiography. From these results we conclude that calcitonin and CGRP secretion in this human C cell model is not differentially affected by alternative RNA processing for the phorbol ester-, and cAMP-dependent secretory pathways.

Calcitonin↗
PubMed1986

Dexamethasone differentially affects the levels of calcitonin and calcitonin gene-related peptide mRNAs expressed in a human medullary thyroid carcinoma cell line.

The TT cells are a continuous line of human C-cells derived from a medullary thyroid carcinoma. These cells produce large quantities of calcitonin and calcitonin gene-related peptide (CGRP) by the differential splicing of a single calcitonin gene transcript. We have used specific cDNA probes for calcitonin and CGRP to study the regulation of the calcitonin gene by dexamethasone, a synthetic glucocorticoid. Northern blot analysis of total cellular RNA isolated from the TT cells showed hybridization of the calcitonin probe to 3600- and 1000-base RNA species. The CGRP probe hybridized to 3600- and 1050-base RNA species. Dexamethasone treatment (10(-9) to 10(-5) M) of TT cells (for 6 days) caused a dosage-dependent increase in calcitonin mRNA levels and a decrease in CGRP mRNA levels. These findings were confirmed in time course studies where dexamethasone treatment (10(-6) M) caused a 2-13-fold increase in calcitonin mRNA and a 40-60% decrease in CGRP mRNA between 4 and 6 days of treatment; the effect was reversible after dexamethasone withdrawal. After excluding an effect of dexamethasone on calcitonin and CGRP mRNA stability, we have concluded that dexamethasone affects the splicing mechanism to favor production of calcitonin mRNA over CGRP mRNA.

Calcitonin↗
PubMed1986

The regulation of somatostatin production in human medullary thyroid carcinoma cells by dexamethasone.

There have been few studies of physiological importance on the regulation of somatostatin by hormones. We have studied the effect of the synthetic glucocorticoid dexamethasone on somatostatin production in the human medullary thyroid carcinoma TT cell line, a model for somatostatin production by the parafollicular cell. Dexamethasone inhibited somatostatin production in a dose-related manner with a maximal effect at a concentration of 10(-6) M. TT cells treated with dexamethasone (10(-6) M) showed an almost complete inhibition of somatostatin peptide production by 48 h of treatment. Molecular sizing chromatography demonstrated a decrease in both the probable somatostatin precursor (13,000 dalton) and the fully processed peptide. Analysis of mRNA content by hybridization revealed that dexamethasone also caused a decrease in detectable somatostatin mRNA. The hybridizable somatostatin mRNA decreased to approximately 50% of basal levels within 12 h of treatment. Northern blot hybridization showed a decrease in a single RNA species representing mature somatostatin mRNA. Dose-response experiments revealed inhibition of both peptide and mRNA at concentrations from 1 X 10(-8) to 1 X 10(-5) M dexamethasone. Four days after withdrawal from dexamethasone treatment, peptide and mRNA levels were higher than dexamethasone-treated controls. The sex steroid estradiol had no inhibitory effect on somatostatin production. These results suggest a potential regulator of somatostatin production and provide a system for the study of somatostatin gene regulation.

Cell Line↗
PubMed1986

Somatostatin production by a human medullary thyroid carcinoma cell line.

Somatostatin (SRIF, SRIF-14) is a known product of the normal and malignant parafollicular cell of the thyroid. In this report we characterize SRIF production by the TT cells, a line of transformed calcitonin-producing cells derived from a human medullary thyroid carcinoma. The cells were found to contain (5-12 ng/10(6) cells) and secrete (3-10 ng/10(6) cells X 48 h) immunoreactive SRIF. Molecular sieve chromatography of cell extracts under denaturing conditions showed a major peak with a mol wt slightly larger than 12,700, probably representing pro-SRIF and a second peak which coeluted with SRIF; in one gel chromatogram a very small peak was also noted which coeluted with SRIF-28, but represented less than 0.4% of the total immunoreactive SRIF. Short term secretion of calcitonin and SRIF was stimulated by calcium in vitro (0.5-4 mM) in a dose-related manner. mRNA isolated from the TT cells hybridized to a specific bovine fetal pancreatic SRIF DNA (BFPS-2); there was no hybridization to identical amounts of mRNA from the atT-20/D16, 3T3, or RINC5F cell lines. In vitro translation of the TT cell mRNA followed by immunoprecipitation and sodium dodecyl sulfate-polyacrylamide gel electrophoresis of the product revealed a single protein band of approximately 13,000 daltons. It was completely abolished when the immunoprecipitation was performed in the presence of excess unlabeled SRIF. Northern transfer of TT cell cytoplasmic RNA and hybridization with FBPS-2 cDNA showed a single hybridizing band with an apparent size of approximately 750 nucleotides. Our observations demonstrate the production of SRIF by a continuous line of human medullary thyroid carcinoma cells and provide a model for studying the biosynthesis and secretion of SRIF in the parafollicular cell.

Animals↗
PubMed1983

Pituitary immunoreactive calcitonin-like material: lack of evidence for cross-reactivity with pro-opiomelanocortin.

In mammals, calcitonin (CT) is synthesized, stored, and secreted by intrathyroidal C cells. Several reports have suggested the presence of immunoreactive CT in the pituitary gland. We have studied the rat pituitary gland using a radioimmunoassay for CT and have also found immunoreactive CT-like material. Assay of extracts of whole rat pituitary glands was performed using a radioimmunoassay for human CT, which gave identical dilution curves with synthetic human CT (hCT), synthetic rat CT (rCT), and mouse and rat thyroid extracts, but not with a variety of other pituitary and hypothalamic peptides. Immunoreactive CT (iCT) content of extracts of whole pituitary glands ranged from 6 to 72 pg/mg wet weight of tissue (60-840 pg/whole pituitary gland), whereas iCT was not measureable (less than 5 pg/mg tissue) in similar extracts of hypothalamus and cerebral cortex. Gel filtration studies of pituitary extracts showed a peak of iCT, which eluted with 125I-rCT and diluted in parallel with rCT. To investigate whether the pituitary iCT was related to pro-opiomelanocortin, extracts of ACTH-producing AtT20/D16 cells from mice, which contain the ACTH precursor in large quantities, were examined and no iCT was found. Immunohistochemical studies of rat pituitary glands with peroxidase-antiperoxidase and immunofluorescent techniques showed positive staining for CT in cells in the pars anterior, but not in the pars intermedia of pars nervosa; this staining was not eliminated when the antiserum was absorbed with CT under conditions that completely obliterated staining of rat thyroid glands. Double staining demonstrated essentially two distinct populations of cells, one positive for CT and another positive for ACTH, with less than 1% of the cells positive for both ACTH and CT. Immunoreactive CT-like material was present in the pituitary glands of rats thyroparathyroidectomized 18 days before they were killed, but was diminished. Biosynthetic labeling in vitro of rat pituitary glands with 3H-leucine showed incorporation into prolactin; there was no incorporation into CT. No in vitro secretion of iCT by whole rat pituitary glands either basally or after high K+ stimulation was observed. We conclude that: (1) a substance that has certain immunologic and size characteristics of CT is present in minute amounts in the pituitary gland of rats; (2) this material is not a part of the ACTH precursor; and (3) positive immunohistochemical staining in pituitary glands may not be specific for authentic CT.

Animals↗
PubMed1983

Clonal strains of rat medullary thyroid carcinoma cells that produce neurotensin and calcitonin. Functional and morphologic studies.

Strains of rat medullary thyroid carcinoma cells were established by two different techniques which were designed to ensure that clonal populations originated from single cells. The clonal strains (44-3C1 and 6-23C6) secreted both immunoreactive calcitonin (CT) and neurotensin (NT) and had growth characteristics that were similar to those of the parent cell lines. Both the 44-3C1 strain and the parent 44-2 cell line consistently produced more CT and NT than the 6-23C6 clonal strain or the parent 6-23 cell line. Peptide secretion in these clonal strains was stimulated by calcium and norepinephrine. Both clonal strains, similar to the parent lines, produced 4 to 12 times more NT than CT. Immunohistochemical studies showed that all cells in both clonal strains stained positively for CT. NT was also present in all cells from both strains with approximately 2% of the cells showing intense staining for this peptide. Ultrastructurally, the cells contained membrane-bound secretory granules which had a mean diameter of 95 nm. Secretory granules were relatively numerous in only 2% of the cells where they tended to be concentrated in cell processes. These studies demonstrate that single rat medullary thyroid carcinoma cells produce both CT and NT and that these strains are useful models for studies of NT and CT biosynthesis and for investigations of the regulation of secretion of two peptides from a single cell type.

Animals↗
PubMed1982

Age-related probability of development of hereditary medullary thyroid carcinoma.

Hereditary medullary thyroid carcinoma is inherited as an autosomal dominant trait; at birth each child of an affected parent has a 50% chance of developing the disease. Measurement of plasma calcitonin concentrations after provocative calcium or pentagastrin stimulation has proved useful in the early diagnosis of this disease. To determine the age-related risk of conversion from a negative to a positive provocative test, 445 members of 11 kindreds were studied with sequential tests. Of 159 family members with a 50% risk at birth of developing medullary thyroid carcinoma 38 converted from a negative to a positive test result (mean age of conversion was 15 years). By means of methods previously described for determining the age-related probability for developing Huntington chorea, we present a method for determining the probability of development of medullary thyroid carcinoma. An individual at risk whose test result was negative had the following probability of converting to a positive test result at a later date: age (years)/probability, 0/0.5; 5/0.49; 10/0.41; 15/0.25; 20/0.16; 25/0.10, 30/0.05; and 35/0. We conclude that hereditary medullary thyroid carcinoma is regularly detectable in the pediatric age group and that screening should begin by age 5 years and be continued at regular intervals until age 35.

Adolescent↗
PubMed1980

Characterization of neurotensin production by a line of rat medullary thyroid carcinoma cells.

The rMTC 6-23 cell line was derived from a calcitonin-producing rat medullary thyroid carcinoma. During characterization of these cells we discovered that they also synthesize and secrete neurotensin (NT), a tridecapeptide originally isolated from the hypothalamus and not previously associated with C cells. Immunoreactive NT (iNT) was measured with two antisera: HC-8, which recognizes the COOH-terminal eight amino acids, and TG-1, which binds the NH2-terminal region of NT. By use of these antisera, 2 M acetic acid extracts of rMTC 6-23 cells were found to contain 0.7--5.0 pmol of iNT per mg of cell protein. Successive fractionation of iNT from cell extracts by gel filtration, ion-exchange chromatograpy, and reverse-phase high-pressure liquid chromatography showed at each step a peak of iNT that was indistinguishable from synthetic NT in its chromatographic behavior. Biologic activity of the iNT was confirmed by demonstrating the characteristic fall in arterial blood pressure in the rat after intravenous injection of material purified from rMTC 6-23 cells. Calcium (0.5--4.0 mM) stimulated release of iNT in a dose-dependent manner; the effect was maximal at 3--4 mM calcium. K+ (50 mM) stimulated release of iNT that was maximal in the presence of 1.0--1.5 mM calcium. Synthesis and secretion of iNT by these cells were shown during a 16-day growth experiment. These results demonstrate that rMTC 6-23 cells contain NT and suggest the possibility of an association between neurotensin and calcitonin.

Animals↗
PubMed1980

Effects of L-dopa and bromocriptine on calcitonin secretion in medullary thyroid carcinoma.

The secretion of calcitonin (CT) by thyroid C cells has been reported to be reduced by dopamine in vitro and by L-dopa in patients with medullary thyroid carcinoma (MTC). These results have suggested that dopa uptake ad decarboxylation to dopamine may play an inhibitory role in the control of CT secretion. We studied the effects of either L-dopa or bromocriptine, a dopamine agonist, on CT secretion in 11 patients with MTC. Seven patients had multiple endocrine adenomatosis type II (MEA II), 1 had MEA III, and 3 had sporadic MTC. Metastatic disease was confirmed in 8, was probable in 2, and was absent in 1. L-Dopa (500 mg orally) was given to 10 patients in 12 trials. Serum obtained serially for CT measurement RIA over a 4-h period showed a fall in 4 instances in 3 patients; there was no decrease in 7 patients. Absorption of L-dopa was confirmed by the finding of the expected rise in serum GH concentration in all nonresponders except 1. Bromocriptine administered in increasing doses (7.5-50 mg/day) for periods from 4-18 weeks in 5 patients with metastatic disease failed to lower the serum CT concentration in any patient. In 2 of 4 cases, pentagastrin-induced CT release was augmented during bromocriptine administration. These results show L-dopa suppression of CT secretion only in a minority of patients with MTC; the lack of response to bromocriptine suggessts that dopamine receptor stimulation by dopaminergic ergots does not inhibit CT secretion in this same group of patients.

Adolescent↗